Nucleic acids such as DNA, are used extensively in the field of molecular biology for research and clinical analyses. Common methods for analyzing DNA are Southern blotting, amplification through methods such as polymerase chain reaction (PCR), and sequencing. Using these methods, differences in DNA sequence are determined to aid in gene identification, population screening, pathogen identification and diagnostic testing. All of these analyses require purified DNA samples as the basis for consistent and valid results.
The analysis and in vitro manipulation of nucleic acids is typically preceded by a nucleic acid isolation step in order to free the nucleic acid from unwanted contaminants which may interfere with subsequent processing procedures. For the vast majority of procedures in both research and diagnostic molecular biology, extracted nucleic acids are required as the first step. In a typical DNA extraction protocol, cells containing the nucleic acid of interest are harvested and lysed.
Cell lysis is a process of releasing materials out of a cell by disrupting the cell membrane, and in particular, a process of extracting intracellular materials from a cell to isolate DNA or RNA before further processing with nucleic acid based methods such as PCR and cloning techniques.
Cell lysis methods through cell rupture can be classified into mechanical methods and non-mechanical methods.
Mechanical methods include ultrasonication, disruption using a homogenizer, pressing using, for example, a French press, etc., decompression, pulverization, etc. Non-mechanical methods include chemical methods, thermal methods, enzymatic methods, etc.
Chemical methods, which are non-mechanical methods, use, for example, an acid, a base, a detergent, a solvent, a chaotropic reagent, etc. Especially, a chemical method using a detergent is widely used. Detergents disrupt a lipid double membrane to release cell contents and lyses membrane protein. Detergents are most commonly used to lyse animal cells. Most detergents denature protein. However, a reagent for cell lysis is separately added, and hence a subsequent process of removing the reagent is required. PCR inhibition may occur, and the process takes a long time.
Enzymatic methods use lysozyme, protease, etc.
Thermal methods include freezing-thawing, heating, osmotic impact, electric impact, etc. For example, cell lysis is achieved by contacting cells with a hot object, such as a hot plate, or by repeating a cycle of freezing to −70° C. and thawing to room temperature.
In US 2006/0141556 cell lysis is performed by raising the temperature of the sample by radiating microwaves onto the sample. An increase in vapour pressure is prevented by adding zwitterionic compounds or ionic liquids to the sample.
There is still need for a fast and effective lysis method.